Deregulation of the apoptotic pathway is responsible for resistance to cancer therapeutics designed to induce cell death. Although there have been many advances in understanding apoptosis, the molecular characteristics involved in resistance or sensitivity to specific apoptotic stimuli are unclear. The identification of genes, which influence the cellular response to apoptosis-based therapeutics will significantly advance the field of cancer cell biology and the rational design of effective anti-cancer strategies. In particular, this research will focus on the tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), which selectively stimulates cell death in cancer cells but not normal cells. However the mechanisms by which normal cells and particular transformed cells obtain resistance to TRAIL remain poorly understood. Our long-term goal is to identify novel gene targets for the design of effective anti-cancer therapeutics. The rationale for the proposed work is that libraries of engineered transcription factors are a unique and powerful tool for identifying genes involved in regulating sensitization or resistance to apoptosis-inducing cancer therapies. The overall objective of this proposal is to identify genes to be targeted for improved apoptosis-based therapeutics. Our hypothesis is that genes identified by libraries of engineered transcription factors (ETFs) will modulate sensitivity and resistance to TRAIL-mediated apoptosis. The objective will be accomplished by testing our hypothesis with the following specific aims: (1) Identify genes that regulate sensitivity to TRAIL-induced apoptosis;(2) Identify genes that regulate TRAIL-mediated suppression of tumor development;(3) Evaluate the role of TRAIL receptor and decoy receptor expression in regulating TRAIL sensitivity. Libraries of ETFs have been recently developed by the sponsor's laboratory to activate or repress the expression of genes associated with complex cellular phenotypes. We will use this tool to identify genes that confer resistance or enhance sensitivity to TRAIL in cancerous and normal cells using an apoptosis-based selection strategy. Additionally, we will use these libraries in combination with a human xenograft tumor model to identify genes that regulate TRAIL activity in vivo. Finally, ETFs will be used to regulate TRAIL receptor expression and elucidate the role of these receptors in selective TRAIL activity. The genes identified in this work will serve as the basis for designing cancer therapies that effectively eliminate cancer cells with minimal effects on normal tissues. These treatments will dramatically enhance the quality of life of cancer patients by maximizing the health of non-cancerous organs during treatment, limiting the need for repeated treatments, and decreasing mortality resulting from ineffective cancer therapy.